Multiple sclerosis (MS) is the most common inflammatory demyelinating disease of the central nervous system (CNS) and is the major cause of non-traumatic neurological disability in young adults in North America. Although the etiology of MS is not clear, its onset is characterized by infiltration of immune cells and the destruction of oligodendrocytes and myelin. After 5-15 years, most patients enter a phase of axonal degeneration and gradual, irreversible neurological decline. There is no cure for MS. Current anti- inflammatory or immunomodulatory therapies are partially effective in delaying the disease, but these therapies do not guarantee functional recovery. Replacement of oligodendrocytes and their subsequent remyelination of demyelinated axons could halt and reverse axonal loss and neurological decline. One approach to increase remyelination is through therapeutic stimulation of endogenous repair. This approach is feasible since generation of new oligodendrocytes and spontaneous remyelination do occur in many MS lesions during early stages of the disease, and even in some chronic lesions, although endogenous remyelination eventually fails. The hypothesis of this proposal is that selected small molecules can be used as remyelination therapeutics to stimulate differentiation of oligodendrocytes and remyelination. Two recent technological developments enable the identification and evaluation of such small molecules. First, an optimized method to produce a highly pure population of primary oligodendrocyte progenitor cells (OPCs) from the mouse that can be used as starting material for screening. Second, a high-content, cell-based phenotypic screening method to automatically score differentiated oligodendrocytes in a medium-throughput manner. This innovative platform will be used to screen a small molecule library to identify hits that can promote OPC differentiation. Putative hits from the primary screen will then be evaluated for their ability to activate the expression of oligodendrocyte-specific markers. Potency, cytotoxicity, cell-type selectivity and interaction with known signaling pathways that regulate oligodendrocyte generation will also be evaluated. The goal of this Phase I project is to identify the best small molecule drug candidate that can promote differentiation of OPCs for subsequent Phase II studies and development. Future Phase II projects will include detailed toxicological studies of the lead compound and in vivo testing in the mouse cuprizone model of demyelination. The long-term goal of this project is to develop therapeutic agents that will promote remyelination to reverse neurological decline in MS patients.